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Sci Rep. 2017 Dec 1;7(1):16789. doi: 10.1038/s41598-017-16881-6.

Conservation of folding and association within a family of spidroin N-terminal domains.

Author information

1
Department of Biotechnology & Biophysics, Julius-Maximilians-University Würzburg, Am Hubland, 97074, Würzburg, Germany.
2
Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge, CB2 0QH, United Kingdom.
3
Department of Biotechnology & Biophysics, Julius-Maximilians-University Würzburg, Am Hubland, 97074, Würzburg, Germany. hannes.neuweiler@uni-wuerzburg.de.

Abstract

Web spiders synthesize silk fibres, nature's toughest biomaterial, through the controlled assembly of fibroin proteins, so-called spidroins. The highly conserved spidroin N-terminal domain (NTD) is a pH-driven self-assembly device that connects spidroins to super-molecules in fibres. The degree to which forces of self-assembly is conserved across spider glands and species is currently unknown because quantitative measures are missing. Here, we report the comparative investigation of spidroin NTDs originating from the major ampullate glands of the spider species Euprosthenops australis, Nephila clavipes, Latrodectus hesperus, and Latrodectus geometricus. We characterized equilibrium thermodynamics and kinetics of folding and self-association using dynamic light scattering, stopped-flow fluorescence and circular dichroism spectroscopy in combination with thermal and chemical denaturation experiments. We found cooperative two-state folding on a sub-millisecond time scale through a late transition state of all four domains. Stability was compromised by repulsive electrostatic forces originating from clustering of point charges on the NTD surface required for function. pH-driven dimerization proceeded with characteristic fast kinetics yielding high affinities. Results showed that energetics and kinetics of NTD self-assembly are highly conserved across spider species despite the different silk mechanical properties and web geometries they produce.

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